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WHAT TO DO WITH YOUR PLATES/CLONES WHEN YOU RECEIVE THEM
IRBD
Clones have been grown on LB agar containing 50µg/ml ampicillin,
these should be placed in a 4°C. fridge.As soon as possible, restreak
the clones onto the same medium and incubate overnight at 37°C to
obtain single colonies.
Plates have been replicated in LB broth containing 8% glycerol and 50µg/ml
ampicillin.
71 plates (96-well format) have been shipped frozen in dry ice and should
be placed in a -70°C freezer on arrival.To make further copies replicate
in the above media and incubate overnight at 37°C.
Plates 1-4 are in pBluescript II SK, plates 5-71 are in pSport I.
IRBE
Clones have been grown on LB agar containing 50µg/ml kanamycin,
these should be placed in a 4°C. fridge.As soon as possible, restreak
the clones onto the same medium and incubate overnight at 37°C to
obtain single colonies.
Plates have been replicated in LB broth containing 8% glycerol and 50µg/ml
kanamycin. 6 plates (96-well format) have been shipped frozen in dry ice
and should be placed in a -70°C freezer on arrival.
To make further copies replicate in the above media and incubate overnight
at 37°C.
All 6 plates are in pZero2 vector.
These frozen stocks can be re-grown for plasmid isolation and purification
or PCR amplification.
Clones - A selection of at least 10 single colonies should be
picked and grown in LB broth containing the appropriate antibiotic + 8
% glycerol for subsequent freezing at -70°C.
These frozen stocks can then be regrown for plasmid isolation and purification.
Streaking out for single colonies is essential in case of contamination
or cross-contamination, or if modifications of the original clone have
taken place as the library has been replicated. If there is a problem,
the more single colonies you streak out, the more likely it is that you
can retrieve the original construct. You can also isolate the original
clone from a contaminated sample by following this procedure.
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